Imidazole derivatives as new potent and selective 20-HETE synthase inhibitors

Article abstract of DOI:10.1016/j.bmcl.2003.11.005

In a previous paper, we reported that an imidazole derivative 1 exhibited a potent inhibitory activity of 20-HETE synthase (1; IC₅₀ value of 5.7 nM), but this compound also exhibited little selectivity for cytochrome P450s (CYPs). We examined some derivatives of imidazole 1 which had an amino group on the side chain, and found that a dimethylaminohexyloxy derivative (3g; IC₅₀ value of 8.8 nM) showed potent and selective inhibitory activity.

Full text of DOI:10.1016/j.bmcl.2003.11.005

Bioorganic & Medicinal Chemistry Letters 14 (2004) 333–336
Imidazole derivatives as new potent and selective 20-HETE
synthase inhibitors
Toshio Nakamura,* Hiroyuki Kakinuma, Hiroki Umemiya, Hideaki Amada,
Noriyuki Miyata, Kazuo Taniguchi, Kagumi Bando and Masakazu Sato*
Medicinal Research Laboratories, Taisho Pharmaceutical Co., Ltd, 1-403 Yoshino-cho, Ohmiya, Saitama 330-8530, Japan
Received 16September 2003; accepted 6November 2003
Abstract—In a previous paper, we reported that an imidazole derivative 1 exhibited a potent inhibitory activity of 20-HETE syn-
thase (1; IC50 value of 5.7 nM), but this compound also exhibited little selectivity for cytochrome P450s (CYPs). We examined some
derivatives of imidazole 1 which had an amino group on the side chain, and found that a dimethylaminohexyloxy derivative (3g;
IC50 value of 8.8 nM) showed potent and selective inhibitory activity.
# 2003 Elsevier Ltd. All rights reserved.
20-Hydroxy-5,8,11,14-eicosatetraenoic acid (20-HETE)
is a major metabolite of arachidonic acid (AA) pro-
duced in the kidney,1 and its biological properties have
recently been extensively studied. The formation of 20-
HETE from AA is catalyzed by cytochrome P450
(CYP) 4A isozymes (CYP4A1, 4A2, 4A3 and 4A8) in
rat kidney2 and cerebral artery,3 and by CYP4A11 and
4F2 in human liver and kidney.4 20-HETE plays an
important role in the regulation of renal vascular and
tubular functions,5ꢀ7 and it also regulates vascular tone
in the cerebral circulation. Therefore, the inhibition of
20-HETE is now considered a promising new target for
the treatment of renal and cerebrovascular diseases. 17-
Octadecynoic acid (17-ODYA),8 1-aminobenzotriazole
(1-ABT),9 N-methylsulfonyl-12,12-dibromododec-11-
enamide (DDMS), 12,12-dibromododec-11-enoic acid
(DBDD)10 and sodium 10-undecynyl sulfate (10-
SUYS)11 are known to inhibit the production of 20-
HETE (Fig. 1). However, these compounds are not
potent and specific inhibitors of 20-HETE formation. In
a previous paper, we reported HET0016( N-hydroxy-N0-
(4-n-butyl-2-methylphenyl)formamidine) as the first
potent and selective 20-HETE synthase inhibitor.12 The
IC50 value of HET0016for the production of 20-HETE
by human renal microsomes was 8.9ꢁ2.7 nM, while its
IC50 value for inhibiting the formation of EETs was
2800 nM.13 HET0016had very little inhibitory activity
for CYP2C9, CYP2D6, CYP3A4 or cyclooxygenase
(COX), even at a higher concentration (100 mM).13
HET0016prevented the acute fall in cerebral blood flow
following subarachnoid hemorrhage (SAH) in the rat.14
Despite its promising pharmacological properties,
HET0016is not soluble enough for injectable formula-
tions under neutral conditions. While the solubility of
HET0016is increased in acidic conditions due to the
basicity of the N-hydroxyformamidine moiety, this is
accompanied by rapid decomposition of the N-
hydroxyformamidine moiety. Recently, we found that
replacement of the N-hydroxyformamidine moiety of
HET0016by an imidazole ring could retain the potent
inhibitory activity of 20-HETE synthase and improve its
acid stability (compound 1; IC50 value of 5.7 nM, Fig.
1).15 However, some imidazole derivatives are known to
show high affinity for the heme iron of major drug-
metabolizing CYP enzymes,16 and 1 also strongly
inhibited the activities of drug-metabolizing CYP
enzymes (CYP 1A2, CYP2C9, CYP2C19, CYP2D6and
CYP3A4; IC50 values of 7, 96, 7, 464 and 348 nM,
respectively).15 Compounds like 1, which strongly inhi-
bit the activities of drug-metabolizing CYP enzymes
may cause unfavorable drug–drug metabolizing inter-
actions. We sought to improve the selectivity of 1
toward drug-metabolizing CYP enzymes by evaluating
the residues on the 1-imidazolylphenyl ring.
Compounds 1 and 3a–g were prepared from 1-(4-
hydroxyphenyl)imidazole 2 by the method shown in
Scheme 1. Reaction of 2 with 1-butanol under Mitsu-
Keywords: 20-HETE.
* Corresponding author. Tel.:+81-48-669-3029; fax:+81-48-652-7254;
e-mail: toshio.nakamura@po.rd.taisho.co.jp
0960-894X/$ - see front matter # 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2003.11.005

334
T. Nakamura et al. / Bioorg. Med. Chem. Lett. 14 (2004) 333–336
nobu conditions afforded the imidazole 3a. Imidazole
derivatives 3b–e were prepared from the corresponding
alcohols under the same conditions as for 3a. Com-
pounds 6a and 6b were prepared according to the
method shown in Scheme 2. Alkylation of 4-fluoro-
nitrobenzene 4 with 3-(2-pyridyl)propanol and sodium
hydride gave 4-[3-(2-pyridyl)propyloxy]nitrobenzene 5a,
and the use of 2-(dimethylamino)ethanol instead of 3-
(2-pyridyl)propanol in the same reaction gave 4-[2-
(dimethylamino)ethoxy]nitrobenzene 5b. The nitro-
benzene derivatives 5a and 5b were treated by the
method reported by Gall et al.17 to give the imidazole
derivatives 6a and 6b. Compound 9 was prepared
according to the method shown in Scheme 3. Treatment
of 2,5-dibromopyridine 7 with n-heptylalcohol and
sodium hydride in N,N-dimethylformamide afforded 5-
bromo-2-heptyloxypyridine 8, successive treatment of
which with imidazole by the method reported by Buch-
wald et al.18 gave the imidazole derivative 9.
alkoxy side chain of 1, which may decrease the affinity
toward drug-metabolizing CYPs. For example, we
introduced some polar groups, like an amino group, to
the side chain of 1. All of the synthesized compounds
were evaluated with regard to their inhibitory activity
toward the human 20-HETE synthesizing enzyme15 and
the major drug-metabolizing CYP enzymes, CYP 1A2,
20
CYP2C9, CYP2C19, CYP2D6and CYP3A4.
The
observed IC50 values are shown in Table 1 along with
the calculated ACD-logD values. Replacement of the
butoxy group of 1 with a pyridin-2-ylmethoxy group did
not affect the inhibitory activity toward 20-HETE syn-
thase (compound 3a; IC50 value of 4.5 nM). While
compound 3a showed better selectivity than 1 toward
CYP1A2 (compounds 1 and 3a; IC50 values of 7.0 and
120 nM, respectively), a similar advantage was not seen
for CYP2C19. While extending the methylene chain of
3a to a propylene group (compounds 6a, 3b and 3c) did
not affect the inhibitory activity toward 20-HETE syn-
thase, it tended to increase the inhibitory activity
toward CYP1A2, probably due to an increase in lipo-
philicity (compounds 3a, 6a, 3b and 3c had ACD-logD
values of 1.99, 2.84, 2.85 and 2.81, respectively).
Replacement of the benzene ring of 1 with a pyridine
ring maintained inhibitory activity toward 20-HETE
synthase (compound 9; IC50 value of 7.7 nM). While
compound 9 showed lower inhibitory activity than 1
toward CYP2C9 (IC50 value of >100,000 nM), it
showed strong inhibitory activities toward CYP1A2 and
2C19. The introduction of a cyclic amino substituent to
the butoxy side chain of 1 maintained the inhibitory
activity toward 20-HETE synthase (compounds 3d, 3e;
IC50 values of 8.6and 16.2 nM, respectively), and gave
lower inhibitory activities toward drug-metabolizing
CYP enzymes. While replacement of the butoxy group
of 1 with a 2-dimethylaminoethoxy group greatly
decreased the inhibitory activities toward the drug-
metabolizing CYP enzymes (compound 6b), and
improved the selectivity toward 20-HETE synthase, the
inhibitory activity toward 20-HETE synthase itself was
decreased to less than 1/60 that of 1 (compound 6b; IC50
value of 387 nM). Changing the dimethylaminoethoxy
side chain of 6b to a dimethylaminopropoxy substituent
increased the inhibitory activity toward 20-HETE syn-
thase (compound 3f; IC50 value of 51 nM). This result
suggests that a longer side chain may be preferable for
potent 20-HETE synthase inhibitory activity. Therefore,
we examined dimethylaminohexyloxy derivative 3g,
which has a longer alkoxy side chain than 3f, and found
that it showed potent inhibitory activity toward 20-
HETE synthase (IC50 value of 8.8 nM). It showed very
little inhibitory activity toward the drug-metabolizing
CYP enzymes (CYP 1A2, CYP2C9, CYP2C19,
Generally, CYP oxidizes lipophilic xenobiotics to intro-
duce a polar substituent so that they can be excreted,
and it has been shown that the increased lipophilicity of
such compounds is associated with an increased cataly-
tic efficiency for CYP.19 Therefore, we examined the
introduction of some polar functional groups to the
Figure 1. Structures of HET0016, 1-ABT, 17-ODYA, DDMS, DBDD
and 10-SUYS.
Scheme 1.
CYP2D6and CYP3A4; IC
values of 410, 4460,
50
Scheme 2. Reagents: (a) ROH, NaH, DMF; (b) H2, Pd/C, MeOH; (c) CH(OEt)3, 100 ꢂC; (d) 1-(2,2-dimethoxy)ethylamine, MeOH, reflux;
(d) TiCl4, DME.

T. Nakamura et al. / Bioorg. Med. Chem. Lett. 14 (2004) 333–336
335
Scheme 3.
Table 1. Inhibition of AA metabolism involving human 20-HETE synthesizing enzyme and inhibitory activities against drug-metabolizing CYPs
by new heterocyclic compounds
Compd
1
RO
X
IC50 (nM)a
P450 inhibition IC50 (nM)b
ACD-LogD
1A2
7.0
2C9
2C19
2D63A4
.8 pH 6
CH
CH
CH
CH
CH
N
5.7
967.00
130
46348
3.38
3a
4.5
120
<461040
20<466
30
6
1.99
2.84
6a
2.1
<46129
<46185
<46101
<46
252
3bc
3c
1.1
<461300
<461040
<461000
399 70
417
344
2.85
1.1
2.81
9c
7.7
>100,000
91
,90076
56
3.74
3d
CH
CH
CH
CH
CH
8.61480
162240
387
1810
0.91
3ec
6bd
3fd
3gc
1440 9
15,000
3400
342600
>100,000
2.27
ꢀ0.08
ꢀ0.41
0.51
40,000
32,800
33,200
2960
9700
6220
1050
>100,000
39,000
4460
51
400
410
2.2
800
a IC50 value for 20-HETE production from AA by human renal microsome.
b IC50 was estimated for each test substance and each enzyme, according to the method of Crespi et al.20
c These compounds were evaluated as the 2HCl salt.
d These compounds were evaluated as the 2 p-toluenesulfonic acid salt.
33,200, 6220 and 800 nM, respectively). It is quite
unique for an imidazole analogue like 3g to have such
selective inhibitory activity toward CYP enzymes.
selectivity without effecting its inhibitory activity
toward 20-HETE synthase. 2-Dimethylaminohexyloxy
derivative 3g exhibited strong inhibitory activity against
20-HETE synthase with an IC50 value of 8.8 nM, and
more than 180-fold to over 15,000-fold selectivity
toward CYP1A2, 2C9, 2C19, 2D6and 3A4.
In conclusion, the introduction of a dialkylamino group
to the butoxy side chain of 1 greatly improved its CYP

336
T. Nakamura et al. / Bioorg. Med. Chem. Lett. 14 (2004) 333–336
11. Xu, F.; Straub, W. O.; Pak, W.; Su, P.; Maier, K. G.; Yu,
References and notes
M.; Roman, R. J.; Ortiz de Montellano, P. R.; Kroetz,
D. L. Am. J. Physiol. Regul. Integr. Comp. Physiol 2002,
283, R710.
1. Imig, J. D.; Zou, A. P.; Stec, D. E.; Harder, D. R.; Falck,
J. R.; Roman, R. J. Am. J. Physiol. 270, R217. 1996.
2. Ito, O.; Alonso-Galicia, M.; Hopp, K. A.; Roman, R. J.
Am. J. Physiol. 274, F395. 1998.
3. Gebremedhin, D.; Lange, R. D.; Lowry, T. F.; Taheri,
M. R.; Birks, E. K.; Hudetz, A. G.; Narayanan, J.; Falck,
J. R.; Okamoto, H.; Roman, R. J.; Nithipatikom, K.;
Campbell, W. B.; Harder, D. R. Circ. Res. 2000, 87, 6 0.
4. Powell, P. K.; Wolf, I.; Jin, R.; Lasker, J. M. J. Pharma-
col. Exp. Ther. 1998, 285, 1327.
5. Omata, K.; Abraham, N. G.; Schwartzman, M. L. Am. J.
Physiol. 262, F591. 1992.
6. Zou, A.-P.; Imig, J. D.; Kaldunski, M.; Ortiz de Mon-
tellano, P. R.; Zhinhua, S.; Roman, R. J. Am. J. Physiol.
1994, 266, F275.
7. Lin, F.; Rios, A.; Falck, J. R.; Belosludtsev, Y.;
Schwartzman, M. L. Am. J. Physiol. 1995, 269, F806.
8. Muerhoff, A. S.; Williams, D. E.; Reich, N. O.; Cajacob,
C. A.; Ortiz de Montellano, P. R.; Masters, B. S. J. Biol.
Chem. 1989, 264, 749.
9. Su, P.; Kaushal, K. M.; Kroetz, D. L. Am. J. Physiol.
1998, 275, R426.
10. Zou, A. P.; Ma, Y. H.; Sui, Z. H.; Ortiz de Montellano,
P. R.; Clark, J. E.; Masters, B. S.; Roman, R. J. J. Phar-
macol. Exp. Ther. 1994, 268, 474.
12. Sato, M.; Ishii, T.; Kobayashi-Matsunaga, Y.; Amada,
H.; Taniguchi, K.; Miyata, N.; Kameo, K. Bioorg. Med.
Chem. Lett. 2001, 11, 2993.
13. Miyata, N.; Taniguchi, K.; Seki, T.; Ishimoto, T.; Sato-
Watanabe, M; Yasuda, Y.; Doi, M.; Kametani, S.;
Tomishima, Y.; Ueki, T.; Sato, M.; Kameo, K. Br. J.
Pharmacol. 2001, 133, 325.
14. Kehl, F.; Cambj-Sapunar, L.; Maier, K. G.; Miyata, N.;
Kametani, S.; Okamoto, H.; Hudetz, A. G.; Schulte,
M. L.; Zagorac, D.; Roman, R. J. Am. J. Physiol. Heart
Circ. Physiol 2002, 282, H1556.
15. Nakamura, T.; Sato, M.; Kakinuma, H.; Miyata, N.;
Taniguchi, K.; Bando, K.; Koda, A.; Kameo, K. J. Med.
Chem., in press.
16. Yamaguchi, H. Nihon Yakuzaishikai Zasshi 1997, 49, 1829.
17. Gall, M.; Kambar, B. V.; Chidester, C. G.; DuChamp,
D. J. J. Heterocycl. Chem. 1988, 25, 1649.
18. Kiyomori, A.; Marcoux, J. F.; Buchwald, S. L. Tetra-
hedron Lett. 1999, 40, 2657.
19. Upthagrove, A. L.; Nelson, W. L. Drug Metab. Dispos.
2001, 29, 1389.
20. Crespi, C. L.; Miller, V. P.; Penman, B. W. Anal. Bio-
chem. 1997, 248, 188.